Adaptor protein-2 exhibits alpha 1 beta 1 or alpha 6 beta 1 integrin-dependent redistribution in rhabdomyosarcoma cells

Biochemistry. 2002 Jun 11;41(23):7232-40. doi: 10.1021/bi011501f.

Abstract

Downregulation of several signaling pathways, such as those stimulated by growth factor receptors, occurs by internalization of signaling receptors through clathrin-coated pits. The first step in internalization or endocytosis is interaction with AP-2, which results in coated pit formation by assembly of clathrin to AP-2. Changes in endocytosis are reflected in the distribution of AP-2 molecules at the cell surface. Integrins are receptors which mediate attachment to the extracellular matrix and also stimulate numerous intracellular signaling pathways; however, it is not known how signaling through integrins is terminated or downregulated. Endocytosis through clathrin-coated pits offers an attractive mechanism for this. This work explores the relationship between AP-2 and beta(1) integrins. RD cells grown for 24 h on collagen or laminin exhibit a redistribution of AP-2 to the cell periphery relative to those grown on fibronectin or polylysine. The total AP-2 protein levels in the cells are unaffected. Blocking alpha(1)beta(1) integrin ligand binding on collagen prevents this redistribution fully. On laminin where alpha(1)beta(1) and alpha(6)beta(1) integrins are engaged, both receptors must be simultaneously blocked to prevent AP-2 redistribution, confirming that the redistribution depends on the specific engagement of the receptors. Immunofluorescence reveals that the majority of alpha(1)beta(1) integrins colocalize with alpha(6)beta(1) integrins in linear structures identified as focal adhesions. A separate fraction of alpha(1)beta(1) integrins colocalize with AP-2 in coated pits. Interestingly, alpha(6)beta(1) integrins are not located in coated pits, demonstrating that integrin colocalization with AP-2 is not necessary to induce redistribution of AP-2.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Proteins, Vesicular Transport
  • Carrier Proteins / metabolism*
  • Cell Adhesion / physiology
  • Clathrin / metabolism*
  • Clathrin-Coated Vesicles / physiology
  • Collagen / metabolism
  • Down-Regulation
  • Endocytosis / physiology
  • Focal Adhesions / physiology
  • Humans
  • Image Enhancement
  • Integrin alpha1beta1
  • Integrin alpha6beta1
  • Integrins / physiology*
  • Laminin / metabolism
  • Membrane Proteins / metabolism*
  • Microscopy, Confocal
  • Rhabdomyosarcoma / metabolism*
  • Rhabdomyosarcoma / pathology
  • Signal Transduction / physiology
  • Tumor Cells, Cultured

Substances

  • Adaptor Proteins, Vesicular Transport
  • Carrier Proteins
  • Clathrin
  • Integrin alpha1beta1
  • Integrin alpha6beta1
  • Integrins
  • Laminin
  • Membrane Proteins
  • Collagen